Human recombinant insulin-like growth factor I. I. Development of a serum-free medium for clonal density assay of growth factors using balb/c 3T3 mouse embryo fibroblasts

1988 ◽  
Vol 24 (11) ◽  
pp. 1099-1106 ◽  
Author(s):  
Terry L. Riss ◽  
Kenneth P. Karey ◽  
B. Daniel Burleigh ◽  
David Farker ◽  
David A. Sirbasku
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Mouse Embryo ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
Factor I ◽  
Mouse Embryo Fibroblasts ◽  
Recombinant Insulin ◽  
Human Recombinant Insulin

scholarly journals Effect of a null mutation of the insulin-like growth factor I receptor gene on growth and transformation of mouse embryo fibroblasts.

1994 ◽  
Vol 14 (6) ◽  
pp. 3604-3612 ◽  
Author(s):  
C Sell ◽  
G Dumenil ◽  
C Deveaud ◽  
M Miura ◽  
D Coppola ◽  
...  
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Soft Agar ◽  
T Antigen ◽  
Insulin Like Growth Factor ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
Factor I ◽  
Growth Factor I

Fibroblast cell lines, designated R- and W cells, were generated, respectively, from mouse embryos homozygous for a targeted disruption of the Igf1r gene, encoding the type 1 insulin-like growth factor receptor, and from their wild-type littermates. W cells grow normally in serum-free medium supplemented with various combinations of purified growth factors, while pre- and postcrisis R- cells cannot grow, as they are arrested before entering the S phase. R- cells are able to grow in 10% serum, albeit more slowly than W cells, and with all phases of the cell cycle being elongated. An activated Ha-ras expressed from a stably transfected plasmid is unable to overcome the inability of R- cells to grow in serum-free medium supplemented with purified clones. Nevertheless, even in the presence of serum, R- cells stably transfected with Ha-ras, alone or in combination with simian virus 40 large T antigen, fail to form colonies in soft agar. Reintroduction into R- cells (or their derivatives) of a plasmid expressing the human insulin-like growth factor I receptor RNA and protein restores their ability to grow with purified growth factors or in soft agar. The signaling pathways participating in cell growth and transformation are discussed on the basis of these results.

Effect of a null mutation of the insulin-like growth factor I receptor gene on growth and transformation of mouse embryo fibroblasts

1994 ◽  
Vol 14 (6) ◽  
pp. 3604-3612
Author(s):  
C Sell ◽  
G Dumenil ◽  
C Deveaud ◽  
M Miura ◽  
D Coppola ◽  
...  
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Soft Agar ◽  
T Antigen ◽  
Insulin Like Growth Factor ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
Factor I ◽  
Growth Factor I

Fibroblast cell lines, designated R- and W cells, were generated, respectively, from mouse embryos homozygous for a targeted disruption of the Igf1r gene, encoding the type 1 insulin-like growth factor receptor, and from their wild-type littermates. W cells grow normally in serum-free medium supplemented with various combinations of purified growth factors, while pre- and postcrisis R- cells cannot grow, as they are arrested before entering the S phase. R- cells are able to grow in 10% serum, albeit more slowly than W cells, and with all phases of the cell cycle being elongated. An activated Ha-ras expressed from a stably transfected plasmid is unable to overcome the inability of R- cells to grow in serum-free medium supplemented with purified clones. Nevertheless, even in the presence of serum, R- cells stably transfected with Ha-ras, alone or in combination with simian virus 40 large T antigen, fail to form colonies in soft agar. Reintroduction into R- cells (or their derivatives) of a plasmid expressing the human insulin-like growth factor I receptor RNA and protein restores their ability to grow with purified growth factors or in soft agar. The signaling pathways participating in cell growth and transformation are discussed on the basis of these results.

Formulation of a complex serum-free medium (CSM) for use in the co-culture of mouse embryos with cells of the female reproductive tract

1991 ◽  
Vol 3 (1) ◽  
pp. 99 ◽  
Author(s):  
D Sakkas ◽  
AO Trounson
Keyword(s):  
Growth Factors ◽  
Growth Factor ◽  
Embryo Culture ◽  
Reproductive Tract ◽  
Culture Media ◽  
Mouse Embryos ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
Cell Numbers

Co-culture of pre-implantation embryos with cells of the reproductive tract requires a medium that is beneficial to both embryos and cells. However, many studies in this area utilize media originally formulated for specific cell lines. In the present study, a complex serum-free medium (CSM) was formulated on the basis of the ionic compositions of existing embryo culture media and mouse oviductal fluid as well as the concentrations of growth factors that appear to benefit mouse embryo development. The study began by investigating the effect of altering the concentrations of K+ ions (0-40 mM) and sulfate ions (0-10 mM) in embryo culture media on the development of 2-cell mouse embryos. Mouse embryos showed improved cell numbers at the blastocyst stage when cultured in 10 mM K+ compared with Whittingham's T6 medium. Embryos were also cultured in T6 supplemented with bovine serum albumin (BSA) containing various concentrations of insulin, insulin-like growth factors I and II, fibroblast growth factor, and epidermal growth factor. Insulin concentrations of 100 ng mL-1 significantly (P less than 0.05) improved the cell numbers of 2-cell embryos cultured to the morulae and blastocyst stages compared with those cultured in T6 + BSA alone. CSM was formulated on the basis of the results of these experiments and was found to support both improved development of 2-cell mouse embryos and the culture of mouse fibroblast and mouse oviduct cells.

scholarly journals Cell-cycle-specific induction of quiescence achieved by limited inhibition of protein synthesis: counteractive effect of addition of purified growth factors

1985 ◽  
Vol 73 (1) ◽  
pp. 375-387
Author(s):  
O. Larsson ◽  
A. Zetterberg ◽  
W. Engstrom
Keyword(s):  
Cell Cycle ◽  
Protein Synthesis ◽  
Growth Factors ◽  
Growth Factor ◽  
Serum Starvation ◽  
Serum Free Medium ◽  
Free Medium ◽  
Serum Free ◽  
G1 Cells ◽  
Synthesis And Degradation

We have previously shown that Swiss 3T3 cells located in the first part of G1 (post-mitotic G1 cells younger than 4.0 h or G1pm cells) were arrested after 9–10 h in the cell cycle by a short (1-8 h) exposure to serum-free medium or by a short (2-4 h) exposure to low doses of the protein synthesis inhibitor cycloheximide (CH). Kinetic data indicate that such G1pm cells rapidly return to G0 during this brief treatment and thereafter require a preparatory period of 8 h before continuing to G1. Cells older than 4 h, i.e. cells in mid or late G1 are already committed to DNA synthesis (presynthesis or G1ps cells). These cells as well as S and G2 cells were consequently unaffected by the brief serum starvation or the brief treatment with cycloheximide. In the present paper we show that the 10-h intermitotic delay that follows a 1–2 h exposure to serum-free medium can be completely counteracted by the presence of any one of the purified growth factors, epidermal growth factor (EGF), insulin or platelet-derived growth factor (PDGF). In contrast, the intermitotic delay following a longer exposure (8 h) to serum-free medium could no longer be counteracted by EGF or insulin. However, PDGF was still active in this respect. Most interestingly, the 12 h gross intermitotic delay induced by a 4h exposure to CH could be efficiently counteracted by EGF, PDGF or insulin. However, this effect on CH-treated cells could be counteracted by the growth factor only in the presence of 10% serum. This indicates the existence of a cooperative effect between PDGF, EGF or insulin and an unidentified serum factor. The effects on the cell cycle time of brief serum starvation and exposure to CH were compared with the effects on rate of protein synthesis and degradation. Although the effects of serum starvation on protein synthesis and degradation were found to be partially normalized by growth factors, we suggest that growth factors prevent cells from leaving the cell cycle by another mechanism and not merely by affecting the level of overall protein accumulation.

scholarly journals Regulation of rat ovarian cell growth and steroid secretion

1980 ◽  
Vol 86 (2) ◽  
pp. 483-489 ◽  
Author(s):  
CC Johnson ◽  
WE Dawson ◽  
JT Turner ◽  
JH Wyche
Keyword(s):  
Growth Factors ◽  
Cell Division ◽  
Growth Factor ◽  
Cell Population ◽  
Serum Free Medium ◽  
Free Medium ◽  
Ovarian Cell ◽  
Serum Free ◽  
Cholesterol Levels ◽  
Progesterone Secretion

A cultured rat ovarian cell line (31 A-F(2)) was used to study the effect of growth factors (epidermal growth factor [EGF] and fibroblast growth factor [FGF]), a survival factor (ovarian growth factor [OGF]), a hormone (insulin), and an iron-binding protein (transferring) on cell proliferation and steroid production under defined culture conditions. EGF and insulin were shown to be mitogenic (half-maximal response at 0.12 nM and 0.11 muM, respectively) for 31A-F(2) cells incubated in serum-free medium. EGF induced up to three doublings in the cell population, whereas insulin induced an average of one cell population doubling. FGF, OGF, and transferrin were found not to have any prominent effect on cell division when incubated individually with 31A-F(2) cells in serum-free medium. However, a combination of EGF, OGF, insulin, and transferrin stimulated cell division to the same approximate extent as cells incubated in the presence of 5 percent fetal calf serum. EGF or insulin did not significantly affect total cell cholesterol levels (relative to cells incubated in serum-free medium) when incubated individually with 31A-F(2) cells. However, cell cholesterol levels were increased by the addition of OGF (250 percent), FGF (370 percent), or a combination of insulin and EGF (320 percent). Progesterone secretion from 31A-F(2) cells was enhanced by EGF (25 percent), FGF (80 percent), and insulin (115 percent). However, the addition of a mitogenic mixture of EGF, OGF, insulin, and transferrin suppressed progesterone secretion 150 percent) below that of control cultures. These studies have permitted us to determine that EGF and insulin are mitogenic factors that are required for the growth of 31A-F(2) cells and that OGF and transferrin are positive cofactors that enhance growth. Also, additional data suggest that cholesterol and progesterone production in 31A-F(2) cells can be regulated by peptide growth factors and the hormone insulin.

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